Youakim A, Hathaway H J, Miller D J, Gong X, Shur B D
Department of Biochemistry and Molecular Biology, University of Texas M.D. Anderson Cancer Center, Houston 77030.
J Cell Biol. 1994 Sep;126(6):1573-83. doi: 10.1083/jcb.126.6.1573.
Sperm surface beta 1,4-galactosyltransferase (GalTase) mediates fertilization in mice by binding to specific O-linked oligosaccharide ligands on the egg coat glycoprotein ZP3. Before binding the egg, sperm GalTase is masked by epididymally derived glycosides that are shed from the sperm surface during capacitation. After binding the egg, sperm-bound oligosaccharides on ZP3 induce the acrosome reaction by receptor aggregation, presumably involving GalTase. In this study, we asked how increasing the levels of sperm surface GalTase would affect sperm-egg interactions using transgenic mice that overexpress GalTase under the control of a heterologous promoter. GalTase expression was elevated in many tissues in adult transgenic animals, including testis. Sperm from transgenic males had approximately six times the wild-type level of surface GalTase protein, which was localized appropriately on the sperm head as revealed by indirect immunofluorescence. As expected, sperm from transgenic mice bound more radiolabeled ZP3 than did wild-type sperm. However, sperm from transgenic animals were relatively unable to bind eggs, as compared to sperm from wild-type animals. The mechanistic basis for the reduced egg-binding ability of transgenic sperm was attributed to alterations in two GalTase-dependent events. First, transgenic sperm that overexpress surface GalTase bound more epididymal glycoside substrates than did sperm from wild-type mice, thus masking GalTase and preventing it from interacting with its zona pellucida ligand. Second, those sperm from transgenic mice that were able to bind the zona pellucida were hypersensitive to ZP3, such that they underwent precocious acrosome reactions and bound to eggs more tenuously than did wild-type sperm. These results demonstrate that sperm-egg binding requires an optimal, rather than maximal, level of surface GalTase expression, since increasing this level decreases sperm reproductive efficiency both before and after egg binding. Although sperm GalTase is required for fertilization by serving as a receptor for the egg zona pellucida, excess surface GalTase is counterproductive to successful sperm-egg binding.
精子表面的β1,4-半乳糖基转移酶（GalTase）通过与卵被糖蛋白ZP3上特定的O-连接寡糖配体结合来介导小鼠的受精过程。在与卵子结合之前，精子GalTase被附睾来源的糖苷所掩盖，这些糖苷在获能过程中从精子表面脱落。与卵子结合后，ZP3上与精子结合的寡糖通过受体聚集诱导顶体反应，推测涉及GalTase。在本研究中，我们利用在异源启动子控制下过表达GalTase的转基因小鼠，探究增加精子表面GalTase水平将如何影响精卵相互作用。在成年转基因动物的许多组织中，包括睾丸，GalTase表达升高。转基因雄性小鼠的精子表面GalTase蛋白水平约为野生型的六倍，间接免疫荧光显示其在精子头部定位正确。正如预期的那样，转基因小鼠的精子比野生型精子结合更多的放射性标记ZP3。然而，与野生型动物的精子相比，转基因动物的精子相对无法结合卵子。转基因精子结合卵子能力降低的机制基础归因于两个GalTase依赖性事件的改变。首先，过表达表面GalTase的转基因精子比野生型小鼠的精子结合更多的附睾糖苷底物，从而掩盖GalTase并阻止其与透明带配体相互作用。其次，那些能够结合透明带的转基因小鼠精子对ZP3高度敏感，以至于它们过早发生顶体反应，并且比野生型精子更松散地结合卵子。这些结果表明，精卵结合需要表面GalTase表达达到最佳水平，而非最高水平，因为增加该水平会降低卵子结合前后精子的生殖效率。尽管精子GalTase作为卵子透明带的受体对受精是必需的，但过量的表面GalTase对成功的精卵结合产生反作用。
